The endomembrane system of all eukaryotic cells consists of a collection of membrane bound compartments. Movement of materials amongst these compartments is achieved by the regulated trafficking of cargo vesicles between compartments. In the process of differentiation, higher cells re-organize their internal membranes in an enormous variety of ways; expanding or specializing certain compartments, distributing them differently in the cell, or carrying vesicles to new destinations. Though these rearrangements are often critical to the function of the differentiated cell, little is known about how these specializations are imposed on the basic pattern of the secretory pathway. Spore formation in yeast involves a cell division that requires a similar organized rearrangement of the secretory apparatus. In this instance, retargetting of secretory vesicles gives rise to a new membrane compartment, the prospore membrane. This membrane arises by the redirection of secretory vesicles away from the plasma membrane to the cell interior. Prospore membrane formation therefore serves as a model for understanding the developmental^ programmed reorganization of cellular membranes. In addition to the retargetting of secretory vesicles, new genetic requirements are imposed on the fusion and trafficking of these vesicles during sporulation. During the initial coalescence of vesicles into a membrane sheet, fusion of the vesicles requires a specialized docking complex, specific fusion proteins, and a lipid modifying enzyme. We are using a combination of genetic and cell biological approaches to understand how these different activities interact to allow the de novo formation of a new membrane compartment. Once an initial membrane sheet is formed, fusion of vesicles to the membrane is controlled by a distinct set of proteins. The basis for this change is also under investigation. Together these studies will provide insight into how the cell modifies its basic architecture during differentiation.